Dongwoo Chae1, Jun Seok Cha1, Kyungnam Kim2,3, Yeajune Cho2,3, Ye Jeong Lee2,3, Ryunha Chang2,3, Yoonyoung Jang2,3, Yeohun Hyun2,3, Keon Hee Lee2,3, and Dongeun Yong2,3
1. Department of Pharmacology, Yonsei university College of medicine, Seoul, Korea. 2. Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, Seoul, Korea. 3. Microbiotix Corporation, Seoul, Korea.
Objectives: The emergence of multi-drug resistant (MDR) pathogens is one of the greatest threats to global health. The pipeline of new antibacterial drugs in clinical development is inadequate and there is a need for alternate therapeutics. Phage therapy is increasingly recognized as a potential candidate to combat antimicrobial resistance, but among many limitations is its poor characterization of pharmacokinetic-pharmacodynamic (PKPD) properties. Here, we applied dynamical modeling to better understand the efficacy of phage therapy targeting carbapenem-resistant P. aeruginosa based on in vitro and in vivo preclinical data.
Methods: Three phage strains – 1304a, 1172, and 1787 – were used to generate preclinical data characterizing phage adsorption rate, one-step growth curve, and lysis kinetics. Untreated bacterial growth measurements were used to fit a bacterial growth model, whose estimated parameters were used for subsequent modeling of phage-induced bacterial lysis. Adsorption rate constant, latent period, burst size, and the fraction of phage-resistant bacteria were estimated as model parameters. The phage-bacteria interaction model was subsequently linked to mice and rat survival data treated with 1304a phages. Mortality hazard was assumed as being proportional to the total bacterial burden. Constant, Weibull, and log-logistic hazard functions were tested to describe the time-varying hazard. Data analysis was performed using R and Python 3, and nonlinear regression analysis was done using Monolix R2021a.
Results: The lytic activity of the three phages increased with a higher multiplicity of infection (MOI). Phage 1304a showed negligible efficacy except when treated with an MOI of 10, while the other two phages caused bacterial lysis from the lowest MOI of 0.0001. The adsorption rate constants of phages 1304a, 1172, and 1787 were estimated as 1.85e-9, 3.0-e-8, and 6.32e-8 mL/h with a burst size of 124. The fraction of resistant bacteria in 1304a and 1172 was estimated to be 0.0091, but that in 1787 was near zero. In concordance with the poor adsorption rate of 1304a, the in vivo efficacy studies that treated 1304a with an MOI of 50 on both mice and rats showed minimal clinical effect. All animals died within 2 days (except one rat in the lowest dose group that survived beyond 5 days) of bacterial infection despite a statistically significant difference in the mice survival rate relative to the control group in the lowest and highest bacterial doses of 9.4e6 and 8.2e7 CFU (p=0.032 and 4.6e-10, respectively). Phage particles of > 1e7 PFU/mL were confirmed in the lung tissues of all dead animals and their titers were proportional to the administered bacterial dose with the highest dose yielding > 1e10 PFU/mL of phage titer. P. aeruginosa were also recovered from the lung, and phage susceptibility testing revealed that 100% of them were resistant to phage 1304a, suggesting that treatment failure was ultimately due to overgrowth of phage-resistant bacteria. A Weibull function with shape parameter 2 reasonably well described the survival curves in both mice and rats and the mortality hazard showed strong a positive correlation with the model-predicted bacterial burden.
Conclusions: This study successfully applied modeling and simulation methodologies to characterize, understand, and aid in the subsequent development of the phage therapy product. In vitro phage properties well predicted the in vivo efficacy, whereby low adsorption rate constant translated to poor efficacy. Our work is expected to serve as a good reference for future modeling applications directed at developing successful phage therapy.
References:
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Reference: PAGE 30 (2022) Abstr 10022 [www.page-meeting.org/?abstract=10022]
Poster: Drug/Disease Modelling - Infection